KR20100135455A - Method for mass production of microbial cellulose and method for mask pack using there of - Google Patents

Abstract

PURPOSE: A method for manufacturing microbial cellulose and method for manufacturing a mask pack of microbial cellulose are provided to quickly produce a large amount of cellulose. CONSTITUTION: A method for manufacturing microbial cellulose comprises: a step of cutlruing Acetobacter xylinum in a liquid medium to obtain culture liquid; a step of culturing Pichia fermentans in a liquid medium to obtain culture liquid; a step of mixing the culture liquids and adding to a spawn medium; a step of culturing spawn in the spawn medium to obtain a spawn culture medium; and a step of collecting cellulose. The medium for culturing spawn contains 3-7%(w/v) of immature fruits, 2-4%(w/v) of yellow sugar and 0.01-0.1%(w/v) of telomerization molasse.

Description

Method for mass production of microbial cellulose and method for mask pack using there of}

The present invention relates to a mass production method of microbial cellulose and a method of manufacturing a mask pack using the same, and more specifically, to obtain acetobacter xylinum culture solution by culturing Acetobacter xylinum in a liquid medium; Culturing Pichia fermentans in a liquid medium to obtain a Peach Permantans culture solution; Preparing a spawn solution for mass cultivation by adding and culturing the acetobacter xyllium culture medium and the Peach permantans culture solution to the spawn preparation medium; The method for producing microbial cellulose comprising the step of adding the mass culture seed solution to the mass culture medium, followed by mass culture to generate cellulose, and then collecting the generated cellulose, and using the microbial cellulose thus prepared. A method for producing a microbial cellulose mask pack.

Bio cellulose produced by bacteria (bio cellulose) is a natural material has no side effects and does not cause environmental pollution, and because cellulose is edible to help the movement of the intestine. In addition, as a functional material, it can be used for packs for skin improvement, burn treatment, and skin trouble treatment, and it is expected to be widely used for other industrial and medical purposes.

Cellulose is the most abundant biopolymer on the planet and economically important in a variety of fields. Trees (more than 50%) and cotton (more than 94%) are made up of cellulose and are the basic substances that make up the cell walls of almost all eukaryotic plants (Microbial cellulose-the natural power to heal wounds.Wojciech Czaja, Alina Krystynowicza , Stanislaw Bieleckia, R. Malcolm Brown Jr. Biomaterials 27 (2006) 145-151.Microbial cellulose: A New Resource for wood, paper, textiles, food and specialty products.copyright by R. Malcolm Brown, Jr. Department of Botany, The University of Texas at Austin, Austin, Texas 78713-7640.

Some bacteria synthesize cellulose as part of glucose metabolism (Role of water-soluble polysaccharides in bacterial cellulose production., Ishida T, Mitarai M, Sugano Y, ShodaM., Biotechnol Bioeng. 2003 Aug 20; 83 (4): 474 These bacteria include some strains of the genus Acetobacter, and these strains produce 7.72 g / l of cellulose after 72 hours of incubation and 0.107 g / l of cellulose per hour (Cultivation of Acetobacter xylium). for bacterial cellulose production in a modified airlift reactor.Hai-Peng Cheng, Pei-Ming Wang, Jech-Wei Chen and Wen-Teng Wu., Buitechnology and Applied Biochemistry., 2002, 35, 125-132.). These bacterial celluloses have different physical properties from plant cellulose and are ultrafine fibers, and are attracting attention because of their excellent physical strength, harmless biodegradability, high water content and transparency. Because of its physical and chemical properties, it has been used as an industrial material for food, medical, and the like.

Acetobacter xylinum is a bacterium that produces the most abundant cellulose in nature. One cell synthesizes 108 glucose into cellulose for 1 hour, which is a pure cellulose that does not contain lignin and hemicellulose, unlike vegetable (Figure 1). It has the advantage of high strength, constant structural stability and light weight.

The range of application is very wide and it is used for food (low calorie ice cream chips, natadi coco, snacks, candy, diet foods, artificial meats, cholesterol-reducing foods, etc.) and medical equipment for wound healing dressing, capsule packaging, artificial skin substrate. It is used for cosmetics such as skin cream, asistent, artificial nail, and manicure, and as an oil cleaner and toxic substance remover for the environment. In addition, there is a possibility of use in various fields, such as the production of disposable tents, the production of diapers, speaker membranes, special paper.

Conventional microbial cellulose has been mass-cultured in the Philippines and the like, but is used as a food, but the cellulose is too strong to be used as a mask pack. In addition, it is difficult to remove unpleasant odors, so they tend to smell even after the mask pack is manufactured, and the skin contact is not soft. Therefore, in order to produce a cellulose exhibiting the appropriate strength to prepare a mask pack, the manufacturer needs to adjust the strength by culturing directly.

The microbial cellulose produced by the conventional method described above is cultivated using coconut only in mass production, so it is difficult to improve the functionality. Moreover, open culture is being produced locally, and even if this seed was obtained and cultured in Korea, mass production was not successful. This is thought to be due to the influence of any microorganisms present in the environment during open culture in the field. Acetobacter xyllium, a strain that produces cellulose There have been many examples of research to produce a large amount of thick cellulose by cultivation with bay, but it is not getting satisfactory results, and it has not succeeded in industrialization.

Therefore, the present inventors have made diligent research efforts to overcome the problems of the prior art as described above, by switching the concept of microbial culture to co-cultivate bacteria and yeast, so that the food chain in the culture solution is induced to open the culture The present invention has been completed, confirming that commercially available thick cellulose can be economically produced and improved in functionality.

Therefore, the main object of the present invention is to provide a mass production method of microbial cellulose capable of producing a large amount of microbial cellulose in a short time and a microbial cellulose mask pack having excellent skin contact and functionality using such microbial cellulose.

According to an aspect of the present invention, the present invention comprises the steps of culturing Acetobacter xylinum in a liquid medium to obtain an acetobacter xylinum broth; Culturing Pichia fermentans in a liquid medium to obtain a Peach Permantans culture solution; Preparing a spawn solution for mass cultivation by adding and culturing the acetobacter xyllium culture medium and the Peach permantans culture solution to the spawn preparation medium; It provides a method for producing microbial cellulose comprising the step of adding the mass culture seed solution to the mass culture medium, followed by mass culture to generate cellulose, and then collecting the generated cellulose.

In the preparation method of the microbial cellulose of the present invention, the acetobacter xylum is preferably cultured at 30 to 34 ° C. for 20 to 30 hours.

In the method for producing microbial cellulose of the present invention, the cultivation of the Peach permantans is preferably incubated for 20 to 30 hours at 25 to 28 ℃.

In the method for producing microbial cellulose of the present invention, the seed solution is added with 1 to 3% (v / v) of acetobacter xyllium culture medium to the seed production medium and incubated at 30 to 34 ° C. for 20 to 30 hours. When 1 to 3% (v / v) of the tooth permantans culture is added and further incubated at 26 to 30 ° C. for 5 to 10 days, it is preferable to use the culture medium in which the microbial cellulose is formed to a thickness of 5 mm or more.

In the method for producing microbial cellulose of the present invention, the mass culture is preferably incubated at 26 to 30 ° C. for 7 to 10 days by adding 0.01 to 0.1% (v / v) of the seed solution to the medium for mass culture. When the thickness of the cellulose becomes 5 to 10 mm, it is better to collect it.

In the method for producing microbial cellulose of the present invention, the seed production medium is 3-7% (w / v) immature fruit juice, 2-4% (w / v) sulfur sugar and 0.01 to 0.1% (w / v) Low molecular weight molasses).

In the method for producing microbial cellulose of the present invention, the medium for culturing the mass is 3-7% (w / v) immature fruit juice, 2-4% (w / v) brown sugar and 0.05-0.2% (w / It is preferable to include the functional material of v).

According to another aspect of the present invention, the present invention provides a microbial cellulose produced by the above method.

According to another aspect of the present invention, the present invention comprises the steps of pretreating the microbial cellulose to obtain a microbial cellulose for mask pack manufacturing; Removing 90 to 95% of the moisture of the microbial cellulose for mask mask manufacturing and molding the molding machine to obtain a molded product; It provides a method for producing a microbial cellulose mask pack, comprising the step of putting the molded article in a wrapping paper and packing in vacuo.

In the method for producing a microbial cellulose mask pack of the present invention, the pretreatment is carried out by repeating the removal of 40 to 60% water after soaking the microbial cellulose in water for 1 to 2 days until the odor of acetic acid is removed. It is preferable to include, and it is more preferable to include 70 to 90% of the water of the microbial cellulose and to soak for 5 to 10 hours in the plant extract at 30 to 50 ℃ to penetrate the functional material in the plant extract.

In the method for producing a microbial cellulose mask pack of the present invention, the essence is glycerin, sodium hyaluronate, propylene glycol, hydrolides collagen, disodium idieti, imidazolidinyl urea, methyl paraben, dipotassium glycidase , Polyquaternium 51, ethanol, Fiji-60 hydrogenate cast oil, tocopheryl acetate or steering flavor.

According to another aspect of the present invention, the present invention provides a microbial cellulose mask pack prepared by the above method.

Hereinafter, the microbial cellulose mass production method of the present invention and a method of manufacturing a mask pack using the same will be described in more detail step by step.

In the present invention, the microbial cellulose is produced using Acetobacter xylinum . The strain may be obtained through a biological resource center certified as a bacterium that produces the most abundant cellulose in nature, or may be purely separated from the microbial cellulose already produced.

Pure separation of acetobacter xyllium from the microbial cellulose can be easily performed by any person of ordinary skill in the art, and in general, by inducing a single colony to be formed by culturing using a solid medium. Pure separation may be used. The solid medium used may be any solid medium capable of incubating acetobacter xyllium, but solids including glucose, yeast extracts, potassium carbonate (CaCO ₃ ) and agar (agar) Preference is given to using a medium. Purely isolated acetobacter xylinum is then cultivated in a liquid medium for use in the preparation of the seed solution, using a liquid medium containing glucose, yeast extracts and potassium carbonate (CaCO ₃ ). It is preferable to incubate for 20 to 30 hours at 30 to 34 ℃.

In addition, Pchia fermentans cocultured to increase the microbial cellulose production efficiency of acetobacter xyllium in the present invention can be easily obtained through a certified bioresource center as a kind of yeast.

When cultivating the Peach permantans, it is preferable to use a medium containing glucose, yeast extracts, peptones, malt extracts, and 20 to 30 ° C at 25 to 28 ° C. It is preferred to incubate for hours.

In order to produce a large amount of microbial cellulose using the above-mentioned strains, it is preferable to first prepare a seed solution which can easily produce microbial cellulose. At this time, as a method for preparing the seed solution, the culture medium of the two strains is added to the seed culture medium, followed by culturing to confirm the production of microbial cellulose, and the use of the culture medium of the seed culture medium for smooth production as a seed solution. good.

As the medium for seed culture, any medium capable of smoothly culturing both strains may be used, but a medium containing immature fruits, brown sugar, and low molecular weight molasses may be used, and immature fruits are ground and filtered. It is preferable to include 3 to 7% (w / v), 2 to 4% (w / v) of sulfur sugar, and 0.01 to 0.1% (w / v) of low molecular weight molasses.

The immature fruit refers to immature fruits, such as apples, preferably used as an extract, and may be processed in any form as long as it can be supplied as a nutrient to the strain.

As a method of culturing the two strains in the seed culture medium, the culture medium of acetobacter xyllium may be added and cultured, and when the acidity is lowered, a method of further culturing by further adding a culture solution of Peachia permantans may be used. At this time, it is preferable to add the culture solution of acetobacter xyllium to be 1 to 3% (v / v), and incubate at 30 to 34 ° C. for 20 to 30 hours after the addition, and the culture solution of Peacha permantans is 1 to 3. It is preferable to add 3% (v / v), and further addition is preferably carried out for 5 to 10 days at 26 to 30 ℃.

When two strains are cultured in the seed culture medium, when microbial cellulose is formed in the upper layer of 5 mm or more, it is preferable to use the lower culture solution as the seed solution.

As a method of producing the microbial cellulose in a large amount after culturing using the above-described seed solution, the microbial cellulose may be collected after culturing by adding the seed solution to the culture medium. The culture medium may be any medium in which both strains can be cultured smoothly, but 3-7% (w / v) immature fruit, 2-4% (w / v) sugar and 0.05 It is preferable to use a medium containing from 0.2% (w / v) functional material, the seed solution is preferably added to 0.01 to 0.1% (v / v), microbial cellulose at 26 to 30 ℃ after addition After culturing to have a thickness of 5 to 10mm, microbial cellulose may be collected.

In the present invention, a mask pack may be manufactured using the microbial cellulose prepared by the method as described above. The microbial cellulose prepared by the above method has a bad smell due to acetic acid and the like, and thus is difficult to use as it is. Therefore, it is good to perform the pretreatment process, in which the pretreatment process may be used independently or in combination with the method of immersion and compression and immersion in the plant extract. As a immersion and compression method, it is preferable to use a method of removing 40 to 60% of water after immersing microbial cellulose in water for 1 to 2 days, and repeatedly performing the process until the odor of acetic acid is removed. In addition, it is preferable to remove 70 to 90% of the moisture of the microbial cellulose and to soak the plant extract at 30 to 50 ° C. for 5 to 10 hours to infiltrate the functional extract in the plant extract.

The plant extract is a substance added according to the purpose of the pack, and may be any substance that is not harmful to the human body and may be useful for skin beauty, but may use plant ingredients such as gardenia, zinnia, red ginseng or pomegranate. Preferably, it is most preferable to use in the form of extract.

As described above, the microbial cellulose that has undergone the pretreatment may be molded to manufacture a mask pack. At this time, the moisture of the microbial cellulose is preferably removed from 90 to 95%, after which it is good to mold using a molding machine.

It is preferable to apply an essence that can enhance the effect as a mask pack after molding the microbial cellulose, wherein the material used for the essence is glycerin, sodium hyaluronate, propylene glycol, hydrolide collagen, disodium identic A, imidazolidinyl urea, methylparaben, dipotassium glycidhazeate, polyquaternium 51, ethanol, fiji-60 hydrogenate cast oil, tocopheryl acetate or steering flavor can be used alone or in combination have.

In the application of the essence as described above, it is preferable to use a method of putting a molded article of microbial cellulose into a wrapping paper, and then vacuum packaging to minimize the deterioration.

Hereinafter, the present invention will be described in more detail with reference to Examples. These embodiments are only for illustrating the present invention, and thus the scope of the present invention is not construed as being limited by these embodiments.

Example 1 Mass Production of Microbial Cellulose

This embodiment was carried out as shown in FIG.

end. Cellulose Production Microbial Strain Isolation and Pure Culture

Aseptically take an appropriate amount of cellulose from microbial cellulose obtained through cultivators in the Philippines, smear it in solid medium (Glucose 100g, Yeast extract 10g, CaCO ₃ 30g, Agar 15g / ℓ) and incubate at 32 ° C for about 1 day. Colonies were taken to purely separate Acetobacter xylinum . Subsequently, colonies formed on the solid medium were inoculated into liquid medium (Glucose 100g, Yeast extract 10g, CaCO ₃ 30g / L) and incubated at 32 ° C. for 1 day.

I. Culture of Yeast Strains

Peachia Permantans (KCTC 7630) purchased from KCTC was plated on a medium consisting of glucose, yeast extracts, peptone, malt extracts, and agar at 28 ° C. After culturing for 1 day, a single colony was isolated, and inoculated in a liquid medium having the same composition except for agar, and then cultured in suspension at 28 ° C. for 1 day.

All. Preparation of mass culture spawn

To prepare a spawn for mass cultivation, a medium [apple (immature fruit) 50 g / l (filtered with a mixer), brown sugar 30 g / l, low molecular weight molasses 0.5 g / l] was prepared, and then acetobacter in 300 ml of medium. Xylnum 5 ml of the liquid culture solution was added and left to incubate at 32 ° C for 1 day.

After 1 day of culture, when the acidity was lowered, the same amount of the culture solution of Peacha permantans was added and cultured at 28 ° C. for about 1 week. When the cellulose was formed 5mm or more in the upper layer was used as a seed for mass culture.

la. Mass Open Cultivation of Cellulose

50g / l of apples (immature fruits), 0.1% of various functional materials (garia, baeknyeoncho, red ginseng or pomegranate extract), 30g / l of brown sugar, 0.5g / l of low-molecular molasses were prepared in large quantities to produce seeds per 20ℓ. 10 ml each was inoculated and cultured in a 28 ° C. incubation chamber (FIG. 3). When the thickness of the cellulose was 5mm or more, it was collected (FIG. 4) and used to prepare a mask pack after post-treatment.

Example 2. Preparation of Cellulose Mask Pack

This embodiment was carried out as shown in FIG.

end. Pretreatment of Cellulose

Cellulose immediately after incubation has a strong acetic acid odor, and if a mask pack is prepared in this state, a slight decay is produced, and thus the commerciality is poor. Therefore, immersion-compression repetition was used to remove acetic acid from the cultured cellulose.

After immersing the cellulose in tap water for 1 day, the method was repeated until the odor of acetic acid was removed by repeating the method of removing water by putting it in a compactor. Finally, about 80% of the water was removed and the plant material (garia, baeknyeoncho, red ginseng or pomegranate extract) in a low temperature state (about 40 ℃ low temperature) was deposited to allow the functional material to penetrate (Fig. 6).

I. Preparation of Essence

Essence was prepared with the components of Table 1 below.

TABLE 1

ingredient content(%) glycerin 5 Sodium hyaluronate 15 Propylene glycol 8 Hydrolides Collagen 5 Disodium ID 0.05 Imidazolidinyl urea 0.15 Methylparaben 0.2 Dipotassium glycidhazeate 0.2 Polyquaternium 51 0.7 Ethanol 2 Fiji-60 Hydrogenate Cast Oil One Tocopheryl acetate 0.2 Steering flavor 0.03 Purified water 62.47

All. Forming and packing

Cellulose sandwich was prepared by spreading the pretreated cellulose on a special pulp and covering the nonwoven fabric (FIG. 7). Twenty of these cellulose sandwiches were stacked and placed in a press to remove 95% moisture. Subsequently, the molding machine was molded (FIG. 8), and then put into a wrapping paper, an essence was added, and vacuum packed.

Example 3. Clinical investigation of microbial cellulose

As a result of surveying 100 women randomly selected for the microbial cellulose mask pack and the general nonwoven mask pack, items such as skin stability, soothing, regenerative effect, elasticity, moisturizing effect, adhesion and dryness, FIG. 9 As shown in the figure, the microbial cellulose mask pack of the present invention was found to be much better than the normal nonwoven pack. In addition, the soft touch felt when the microbial cellulose mask pack is attached is different from other mask packs. In addition, in the case of the normal non-woven mask pack after 30 minutes to dry it is impossible to use, but the microbial cellulose mask pack has the advantage that can be used for more than 12 hours it was confirmed that the effect is much better.

As described above, according to the present invention, when the cellulose producing strain Acetobacter xylinum and the yeast Pchia fermentans are co-cultured using the medium and the culturing method of the present invention, the food is fed into the culture medium. The chain is easy to produce a large amount of cellulose in a short time, the strength of the cellulose can be adjusted to produce cellulose of the appropriate strength according to the purpose, using a mask pack (mask pack) using the produced cellulose When manufacturing, it is possible to manufacture a mask pack excellent in skin contact and functionality.

1 is a diagram illustrating the process of producing cellulose from Acetobacter xylinum .

2 is a flowchart illustrating a production process of microbial cellulose.

Figure 3 is a photograph showing a state of culturing in a 28 ℃ culture chamber using a 30 cm × 20 cm culture vessel during the mass production of microbial cellulose.

Figure 4 is a photograph showing the state of collecting the microbial cellulose produced by the mass production method.

5 is a flowchart illustrating a process of producing a microbial cellulose mask pack.

6 is a photograph showing a state after the deposition of a plant extract of microbial cellulose.

Figure 7 is a photograph showing the appearance of microbial cellulose attached with a nonwoven fabric and pulp.

Figure 8 is a photograph showing the appearance of the molded product after the formation of microbial cellulose.

9 is a graph showing a survey result for the microbial cellulose mask pack.

Claims (13)

a) culturing Acetobacter xylinum in a liquid medium to obtain acetobacter xylinum broth; b) culturing Pichia fermentans in a liquid medium to obtain a Peach Permantans culture solution; c) preparing a seed culture solution for mass cultivation by adding the acetobacter xylinum culture solution and the Peach permantans culture solution to the seed production medium; d) a method for producing microbial cellulose, comprising the step of adding the seed culture solution for mass culture to the medium for mass culture, followed by mass culturing to generate cellulose, and then collecting the cellulose produced. According to claim 1, wherein the step a) is a method for producing microbial cellulose, characterized in that the step of incubation for 20 to 30 hours at 30 to 34 ℃. According to claim 1, wherein the step b) is a method for producing microbial cellulose, characterized in that the step of incubation for 20 to 30 hours at 26 to 30 ℃. According to claim 1, wherein the step c) is added 1 to 3% (v / v) of acetobacter xyllium culture medium in the spawn preparation medium and incubated for 20 to 30 hours at 30 to 34 ℃, Peach permantans Method for producing microbial cellulose, characterized in that the step of adding a culture solution 1 to 3% (v / v) and further culture for 5 to 10 days at 26 to 30 ℃ to produce a spawn for mass culture. The method of claim 1, wherein d) is added to the culture medium for the mass culture medium to 0.01 to 0.1% (v / v) incubated for 5 to 10 days at 26 to 30 ℃, the thickness of the resulting cellulose is 5 to Method for producing microbial cellulose, characterized in that the step of collecting when the 10mm. According to claim 1, wherein the seed production medium is 3-7% (w / v) of immature fruit, 2 to 4% (w / v) sugar and 0.01 to 0.1% (w / v) low molecular weight molasses Method for producing a microbial cellulose, characterized in that the medium containing. The method of claim 1, wherein the mass culture medium is 3-7% (w / v) immature fruit, 2 to 4% (w / v) sugar and 0.05 to 0.2% (w / v) functional material Method for producing a microbial cellulose, characterized in that the medium containing. Microbial cellulose produced by the method of any one of claims 1 to 7. a) pre-treating the microbial cellulose of claim 8 to obtain microbial cellulose for mask pack manufacturing; b) removing the water of the microbial cellulose for mask mask manufacturing by 90 to 95% and molding a molding machine to obtain a molding; c) The method of manufacturing a microbial cellulose mask pack comprising the step of putting the molded article in a wrapping paper and putting the essence in vacuum. 10. The method of claim 9, wherein the pretreatment in step a) comprises submerging microbial cellulose in water for 1 to 2 days and then removing 40 to 60% water until the acetic acid odor is removed. Method for producing a microbial cellulose mask pack, characterized in that. 10. The method according to claim 9, wherein the pretreatment in step a) removes 70 to 90% of the moisture of the microbial cellulose and soaks the plant extract at 30 to 50 ° C. for 5 to 10 hours to allow the functional material in the plant extract to penetrate. Method for producing a microbial cellulose mask pack, characterized in that it comprises a. 10. The method of claim 9, wherein the essence is glycerin, sodium hyaluronate, propylene glycol, hydrolides collagen, disodium id, imidazolidinyl urea, methylparaben, dipotassium glycidhazeate, polyquaternium 51 Method for producing a microbial cellulose mask pack, characterized in that at least one selected from the group consisting of, ethanol, Fiji-60 hydrogenate cast oil, tocopheryl acetate and steering flavor. A microbial cellulose mask pack prepared by the method of any one of claims 9 to 12.

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